This proposal seeks to continue to investigate the neurochemical basis of ethanol-seeking behavior, with a shift in emphasis toward the investigation of the neurobiological basis of relapse. Alcoholism is a chronic relapsing disorder and vulnerability to relapse following withdrawal presents a great challenge for the treatment of alcohol addiction. The clinical literature suggests that one of the primary factors underlying high rates of relapse in detoxified alcoholics are conditioned responses elicited by ethanol-related environmental stimuli that lead to craving or the compulsion to consume ethanol. Yet, experimental studies of cue-induced alcohol-seeking behavior and its neurobiological basis are rare. Moreover, pertinent information is restricted to nondependent rats and, thus, provides only a limited heuristic basis for the understanding of the stimulus control of ethanol-seeking behavior and its neurobiological substrates in human alcoholics. The objective of this proposal is to investigate experimentally the role of ethanol-associated environmental stimuli in the initiation of ethanol-seeking behavior after abstinence, and to identify neuroanatomical, neurochemical, and neuropharmacological substrates for these conditioned effects of ethanol cues. Specific Aim 1 will utilize an operant response-reinstatement model of relapse developed during the previous funding period to establish the time course and resistance to extinction to the behavioral effects of ethanol-associated environmental stimuli, and to compare their effects in nondependent rats to those in rats with a history of dependence. Specific Aim 2 will focus on the identification of neurobiological substrates of cue-induced reinstatement of ethanol-seeking behavior by examining the effects of ethanol cues on neural activation within brain reward regions using Fos immunohistochemistry and by characterizing the effects of ethanol cues on extracellular dopamine (DA) levels within brain reward regions as well as on the release of the stress-regulatory neuropeptide corticotrophin-releasing factor (CRF) with the central nucleus of the amygdala. Experiments in Specific Aim 3 will examine whether the response-reinstating actions of ethanol cues are sensitive to pharmacological manipulation, and to further isolate critical brain regions by combining tests of the effects of ligands for opioid, dopamine, and serotonin receptors on ethanol-seeking behavior with measurements of alterations in cue-induced Fos expression by these agents. By increasing understanding of the neurobiological basis of relapse, these studies are expected to have direct implications for the development of pharmacotherapeutic treatments for the treatment and prevention of compulsive ethanol-seeking behavior and relapse.